Overview

Some customers have reported successful staining using this antibody on frozen sections, however ab15690 is not routinely tested on frozen sections and we do not recommend this antibody for use with frozen tissue.
In our hands this product worked well in WB under standard reducing and denaturing conditions (please see image below). ab15690 detected Iba1 migrating at approximately 17 kDa in positive control samples (Rat spleen, THP1, C6) and bound no protein in samples we believe have little or no Iba1 expression (HEK293, NIH3T3 and A431). We recommend blocking in milk. Blocking with BSA gives high background.

Target

Function

Actin-binding protein that enhances membrane ruffling and RAC activation. Enhances the actin-bundling activity of LCP1. Binds calcium. Plays a role in RAC signaling and in phagocytosis. May play a role in macrophage activation and function. Promotes the proliferation of vascular smooth muscle cells and of T-lymphocytes. Enhances lymphocyte migration. Plays a role in vascular inflammation.

Images

IHC image of Iba1 staining in human normal spleen formalin fixed paraffin embedded tissue section*, performed on a Leica Bond™ system using the standard protocol F. The section was pre-treated using heat mediated antigen retrieval with sodium citrate buffer (pH6, epitope retrieval solution 1) for 20 mins. The section was then incubated with ab15690, 10µg/ml, for 15 mins at room temperature and detected using an HRP conjugated compact polymer system. DAB was used as the chromogen. The section was then counterstained with haematoxylin and mounted with DPX.

This blot was produced using a 4-12% Bis-Tris gel under the MOPS buffer system. The gel was run at 200V for 60 minutes before being transferred onto a nitrocellulose membrane at 30V for 70 minutes. The membrane was then blocked for an hour before being incubated with ab15690 (anti-Iba1 antibody; 1 ug/mL) for 18 hours at 4°C. Antibody binding was detected using ab97040 (HRP-labelled goat anti-mouse IgG) at 1:50,000 dilution for 1 hour at room temperature and visualised using ECL development solution ab133406.

This concentration is indeed quite high for an antibody so if you wish to dilute it in PBS then that is fine. I would say, that diluting it down to 1 mg/ml would be fine and is standard for most antibodies. Diluting it down to 100 ug/ml is quite a big dilution and I would not recommend this as it may affect the stability of this product especially when storing it at -20ºC or -80ºC.I notice that you are wanting to use this antibody for IHC-FrFl on rat brain tissue. As stated on the datasheet, although some customers have reported successful staining using this antibody on frozen sections, we do not routinely test this antibody on frozen sections, and we do not recommend it for use on these. For this reason, we have not suggested a dilution factor for this application. If you still want to proceed with this experiment I would suggest following the IHC-P guidelines so using 10-20 ug/ml of antibody per experiment.

I would like to reassure you that our antibodies have all been tested successfully and would be covered by our guarantee in the applications listed on the datasheets. Therefore, any antibody tested in the species and application you are using should be suitable for your applications.

Also, although we produce many in-house antibodies, we obtain other antibodies from a wide range of sources. Therefore, comparison experiments will not often have been done.

If you wish to be more certain, I can suggest choosing the antibody that has been best characterized. By this I mean the antibody that has been tested in the most applications and has the most data available (on the datasheet).

In regard to your concerns about ab15690 Anti-Iba1 antibody [1022-5], we do offer another Anti-Iba1 which has been raised in goat and has a number of referenced publications, Abreviews and support data. I would recommend this product as an alternative to ab15690.

https://www.abcam.com/iba1-antibody-ab5076.html

I look forward to hearing from you. Should you have any further questions please do not hesitate to ask.

For UNTESTED species and/or applications, we have established a testing discount program. Here is a brief description of how it works:

The testing discount program is for customers who like to use an antibody/protein/kit on an untested species/application. You would purchase the antibody/protein/kit at full price, test it and submit an Abreview with your data (positive or negative). On your next order you will receive a discount for ONE antibody/protein at the full price (100%) of the antibody/protein you have tested, or a full price discount for the amount paid for the kit. The terms and conditions applicable to this offer can be found here: https://www.abcam.com/collaborationdiscount. This programapplies to ELISA kits and other kits we offer.

Also, I wanted to check homology betweenhuman and rat MMP1, but was not able to find the full length protein sequence for rat MMP1 in the database. However, if you like to try any of the antibodies that do not list "rat" as tested species (only as predicted), please let me know and I'd be happy to send you a testing discount code.

I hope this information is helpful to you. Please do not hesitate to contact us if you need any more advice or information.